Cytokines are a group of proteins that cells release upon excitation (only very few cytokines are expressed on cell membranes). Cytokines produced by cells can affect target cells nearby or through blood circulation at very low concentration. They have broad functions on promoting growth, differentiation and activation of target cells. Many cytokines can target immune cells and play a role in immune response. Based on structural and functional differences, cytokines may be broadly divided into chemokines, interleukins, growth factors, transforming growth factors, colony stimulating factors, tumor necrosis factors, and interferons, etc.
Chemokines are a group of cytokines being able to attract leukocytes, which are generally positively charged, secretory proteins having small molecule weights. Their main function is to attract immune cells to a region having tissue injuries or pathogen infection, allowing leukocytes to subsequently perform phagocytosis or elicit inflammation against pathogens at this specific site. Leukocytes attracted by chemokines may include neutrophils, monocytes/macrophages, natural killer cells, dendritic cells and other leukocytes, which are of innate immunity; and T lymphocytes (T cells) or B lymphocytes (B cells) of adaptive immunity. Accordingly, chemokines play a very important role in the immune system of living organisms. Most chemokines have four highly conserved cysteine (C) forming disulfide bonds to stable their structure. Based on different numbers of amino acids between the first two Cs and the procession of the first C or not, they may be classified into four subfamilies of CXC (or α), CC (or β), C (or γ) and CX3C. Stromal cell-derived factor-1 (SDF-1) is classified into the CXC subfamily of chemokines, and is also known as CXC ligand 12 (CXCL12). Having been observed in many species including humans, mice and cats of mammals and Xenopus of amphibians, and zebra fishes, it has little variation between different species and is highly conserved (Shirozu et al., Genomics 28, 495-500). mRNAs transcribed from SDF-1 gene in mice and humans are subject to different splicings and thus two isoforms of SDF-1 may be observed: SDF-1α and SDF-1β. The distribution of SDF-1 is very wide, and can be detected, including in lymphoid tissue, kidney, lung, liver, brain and muscle (Shirozu et al., Genomics 28, 495-500). SDF-1 receptor CXCR4 not only constantly presents in organs, but can also be seen in hematopoietic stem cells, endothelial cells, dendritic cells, B cells and T cells. Therefore, these cells are attracted by SDF-1 to migrate to the site with high concentration of the chemokines (Bleul et al., Nature, 382: 829-833; Oberlin et al., Nature 382: 833-835; Read et al., Developmental and comparative immunology, 29, 143-152). Interleukin-8 (IL-8) is also classified into the CXC subfamily of chemokines (also known as CXCL8). After initial discovery in humans, it was successively observed in economic animals of pigs, cows and chickens. IL-8 at low concentration is able to attract several immune cells, including monocytes, macrophages, lymphocytes, neutrophils, etc.
CD40 ligand (CD40L) is a member of tumor necrosis factor (TNF) superfamily, which is a cytokine having functions on tumor necrosis and promoting differentiation, proliferation and apoptosis of white blood cells. CD40L is synthesized as a transmembrane protein. Take human CD40L as an example, the protein has a total of 261 amino acids, with first 22 amino-terminal amino acids being intracellular region, followed by 24 amino acids being transmembrane region, and 215 carboxy-terminal amino acids being extracellular (Exc) region, wherein the Exc region has at its carboxy terminus a TNF homology (TNFh) region conserved for all TNF superfamily proteins. CD40L presents mainly in the form of a transmembrane protein on the surface of activated CD4+ T cells, and also presents on CD8+ T cells, basophils, eosinophils, mast cells, natural killer cells, platelets, and even on the surface of CD40-expressing cells.
CD40, receptor of CD40L, is distributed on the surfaces of antigen presenting cells (APCs) of B cells, dendritic cells, macrophages, etc. Physiologically, these antigen presenting cells can be activated by CD40L expressed by T helper cells, promoting the expression of major histocompatibility complex class II (MHC-II) molecules and B7 molecules to assist in antigen presentation. CD40L activates signal transduction pathways by binding to CD40 on target cells. In addition to the aforementioned promotion of antigen presentation, effecting on B cells, CD40L can promote B cell proliferation, isotype switching of immunoglobulins, antibody secretion, memory B cell differentiation, or prevention of apoptosis; effecting on macrophages, CD40L can enhance their activation, production of interleukin-12 (IL-12) to activate T helper 1 (Th1), or secretion of chemokines, or the production of nitric oxide (NO) to promote microorganism defense ability of macrophages; effecting on dendritic cells, it can make them mature and activated, wherein the mature dendritic cells not only express a large amount of MHC-II molecules to promote antigen presentation, but also secrete chemokines of TNF-α and IL-8, macrophage inflammatory protein 1a (MIP-1a), etc.
There are many researches that apply CD40L on vaccine adjuvant or treatment, for example, as adjuvants for duck hepatitis B virus (DHBV) vaccines (Gares et al., Clin Vaccine Immunol 13, 958-965), human immunodeficiency virus (HIV) DNA vaccines (Stone et al., J Virol 80, 1762-1772), or in the treatment of human autoimmune diseases (Howard & Miller, Autoimmunity 37, 411-418), etc.
IL-2 is classified into the hematopoietin family, the family including a number of cell growth-related hormones or other cytokines, etc. Functions of IL-2 include: regulating the maturation and differentiation of T cells, stimulating proliferation and antibody secretion of B cells, promoting cytotoxicity of natural killer cells, and activating monocytes and macrophages, etc. IL-2 can also stimulate T cells and B cells to continue expressing MHC, and also stimulate natural killer cells to produce several different cytokines, including TNF-α, IFN-γ and granulocyte/macrophage colony stimulating factor (GM-CSF), etc. Studies have shown that IL-2 has anti-tumor and vaccine-enhancing effects.
However, there remains a need in the art for cytokines and chemokines with an improved activity.